Ovarian steroid hormones, estrogen (E) and progesterone (P), exert profound influence on development, reproduction, and aging through activation of cognate nuclear receptors (ERa, PR isoforms a and b). In addition to synthesis of PR in female hypothalamic ventromedial nucleus (VMN), E induces changes in local microcircuitry, an effect that requires cAMP activity and for receptivity. Since cAMP increases synthesis and release of constitutive pituitary adenylyl cyclase-activating polypeptide (PACAP) in other tissue for local binding to PACAP receptors, the goal of this proposal is to determine whether E-induced PACAP mediates a novel autocrine/paracrine trans-synaptic loop within VMN microcircuitry required for onset of P-facilitated receptivity. Since studies on the biological role of PR have been hampered by a lack of knowledge of the cell-lineages that express PR isoforms, mice with selective ablation of PRa (PRAKO) and PRb (PRBKO) will be studied. The specific aims of this project are: 1) To determine whether E and P and their cognate receptors [ERa, PRa, PRb] regulate the synthesis and release of pituitary adenylyl cyclase-activating polypeptide (PACAP) in the microcircuitry of the VMN ultimately for receptivity, 2) To characterize the gene expression profile ('molecular fingerprints') of individual PR-expressing and adjacent nonPR- expressing cells in the VMN that mediate isoform-specific PR- facilitated behavior, 3) To identify the steroid-induced mechanism(s) and its major components (VMN expression pattern of membrane-bound receptors [PAC1, VPAC, VPAC2]; signaling pathway [cAMP, MAPK] by which PACAP mediates isoform-specific PR- facilitated behavior. Well-established procedures [in situ hybridization, blot analysis, immunohistochemistry, steroid receptor-dependent behavioral] and new technologies [real-time RT-PCR, single cell expression profiling (molecular fingerprinting), microarrays, Laser capture of individual cells] will be used. By monitoring coordinate changes in PACAP activity, intracellular signaling, and gene expression induced specific steroid receptors (PRa and PRb) in the individual VMN cells that correlate with hormone status and behavior, unique insight into the biological consequences of expressed genetic variability of cells will be identified. This will furnish insight into the underlying cellular and molecular mechanisms that produce steroid receptor-dependent disease etiologies and should contribute to the development of new therapeutic strategies.